[Multidisciplinary regenerative treatment and mechanisms for rescuing a severe calciphylaxis patient with human amnion-derived mesenchymal stem cells].

Calciphylaxis is a rare disease with severe pain and high-mortality due to cutaneous ischemic necrosis and infection that currently lacks proved effective therapies. The occurrence of calciphylaxis in end stage kidney disease (ESKD) patients is known as calcific uremic arteriolopathy (CUA), which is characterized histologically by dermal microvessel calcification, intimal fibroplasia and microthrombosis. Here we innovatively treated a severe CUA patient with human amnion-derived mesenchymal stem cells (hAMSCs). A 34-year-old uremic woman was presented with progressive, painful malodorous ulcers in buttocks and mummified lower limbs. Skin pathological features supported the diagnosis of calciphylaxis. The patient was refractory to conventional multidisciplinary symptomatic therapies. With the approval of our hospital ethics committee, she was treated with hAMSCs including intravenous and local intramuscular injection, and external application of hAMSC culture supernatant to the wound area. During 15-month follow-up, the patient had regeneration of skin and soft tissues, with improved blood biochemical, inflammatory, mineral and bone metabolic indices and immunoregulation effects. After 15-month hAMSC treatment, the score of pain visual analog scale (VAS) decreased from 10 to 0, Bates-Jensen wound assessment tool (BWAT) score decreased from 65 to 13, and wound-quality of life (Wound-QoL) questionnaire score decreased from 68 to 0. We propose that hAMSC treatment is promising for CUA patients. The therapy is potentially involved in the multiple beneficial effects of inhibiting vascular calcification, stimulating angiogenesis and myogenesis, modulating adverse inflammatory and immunologic responses, promoting re-epithelialization and restoring skin integrity.

[Related factors for hyperkalemia and its recurrence in maintenance hemodialysis patients].

Objective: To investigate the prevalence of hyperkalemia and recurrent hyperkalemia in maintenance hemodialysis patients, and identify the related factors. Methods: The general information and clinical data of maintenance hemodialysis patients in the hemodialysis center of the First Affiliated Hospital of Nanjing Medical University from January to December 2020 were collected. According to the level of serum potassium, the patients were divided into normokalemia group and hyperkalemia group. Patients in the hyperkalemia group were further divided into single hyperkalemia group and recurrent hyperkalemia group according to the number of occurrences of hyperkalemia. Then, the prevalence of different serum potassium abnormalities and the related factors were explored. Results: Among the 352 patients included in the final analysis, 129 cases (36.6%) were in the normokalemia group [mean age: (62±15) years, 99 males] and 223 cases (63.4%) were in the hyperkalemia group [mean age: (60±14) years, 153 males]. Multivariate logistic regression analysis demonstrated that, compared with the normokalemia group, night-time dialysis (OR=4.012, 95%CI: 1.519-10.601, P=0.005), concurrent diabetes (OR=1.947, 95%CI: 1.148-3.304, P=0.013) and the number of serum potassium tested before hemodialysis (OR=1.561, 95%CI:1.292-1.885, P<0.001) were independent risk factors for hyperkalemia. Among 223 patients with hyperkalemia, 78 cases (35.0%) were in single hyperkalemia group and 145 cases (65.0%) were in recurrent hyperkalemia group. Multivariate logistic regression analysis demonstrated that, serum calcium (OR=21.885, 95%CI: 3.740-128.077, P=0.001), peak value of serum potassium before hemodialysis (OR=63.157, 95%CI: 25.265-157.876, P<0.001) and the number of serum potassium tested before hemodialysis (OR=1.814, 95%CI: 1.378-2.388, P<0.001) were the independent risk factors for the recurrence of hyperkalemia. Conclusions: The prevalence of hyperkalemia is high in maintenance hemodialysis patients, especially in those with diabetes or night-time dialysis. It is necessary to monitor serum potassium regularly. In addition, high serum calcium and peak value of serum potassium before hemodialysis are related factors for recurrent hyperkalemia.

[Association of urinary sodium excretion with fluid overload in patients with chronic kidney disease].

Objective: To investigate the relationship between urinary sodium excretion and fluid overload (FO) in non-dialysis patients with chronic kidney disease (CKD). Methods: Patients with CKD stage 1-4 who underwent bioelectrical impedance (BIA) in the Department of Nephrology, Jiangsu Province Hospital from December 2019 to January 2021 were recruited. All enrolled patients were categorized into two groups according to whether or not they develop FO. Further, clinical parameters were compared between the two groups. Spearman correlation analysis was used to investigate the association between over hydration/extracellular water (OH/ECW) and clinical characteristics. Multivariate logistic regression analysis was performed to evaluate the relationship between urinary sodium excretion and FO (FO was defined as OH/ECW≥7%). Results: A total of 385 patients with CKD stage 1-4 were finally included in the study, with a mean age of (46±15) years. There were 216 male cases (56.1%), and 150 cases (39.0%) existed FO. Spearman correlation analysis indicated that OH/ECW positively correlated with urinary sodium excretion (r=0.147, P=0.004), urinary protein excretion (r=0.555, P<0.001) and systolic blood pressure (r=0.241, P<0.001), but inversely related to estimated glomerular filtration rate (eGFR) (r=-0.111, P=0.030) and serum albumin (r=-0.659, P<0.001). After adjusting for confounding factors including age, systolic blood pressure, diabetes, urinary protein excretion, serum albumin, serum sodium, serum chlorine, urinary calcium excretion, urinary phosphorus excretion and use of diuretics, multivariate logistic regression analysis demonstrated that higher level of urinary sodium excretion was associated with increased risk of FO in patients with CKD (OR=1.005, 95%CI: 1.000-1.011, P=0.048). Conclusion: High urinary sodium excretion is independently associated with fluid FO in non-dialysis patients with CKD.

[Development of a hyperkalemia risk assessment model for patients with chronic kidney disease].

Objective: To investigate risk factors for hyperkalemia among chronic kidney disease (CKD) patients and establish a risk assessment model for predicting hyperkalemia events. Methods: Clinical data of CKD patients (stage 3 to 5) hospitalized between May 2017 and June 2020 from 14 hospitals were retrospectively collected and divided into training dataset and validation dataset through balanced random sampling. Multivariate logistic regression analysis was used to analyze risk factors for hyperkalemia in CKD patients and the factors were scored. Receiver operating characteristic (ROC) curve was plotted and the area under the curve (AUC) was calculated. Meanwhile, the cut-off value with the best sensitivity and specificity were used to verify the accuracy of the model in validation dataset. Results: A total of 847 CKD patients were enrolled and further divided into training dataset (n=675) and validation dataset (n=172). There were 555 males and 292 females, with a mean age of (57.2±15.6) years. Multivariate logistic regression analysis showed that age, CKD stage, history of heart failure, history of serum potassium ≥5.0 mmol/L, diabetes, metabolic acidosis, and use of medications that increase serum potassium levels were risk factors for causing hyperkalemia in patients with CKD. Risk assessment model was established based on these risk factors. The AUC of the ROC curve was 0.809. Using 4 as the cut-off value, the sensitivity and specificity for predicting hyperkalemia events reached 87.1% and 57.0%, respectively. Conclusion: The model established in the current study can be used for predicting hyperkalemia events in clinical practices, which offers a new way to optimize serum potassium management in patients with CKD.

Plasma FGF23 levels and heart rate variability in patients with stage 5 CKD.

UNLABELLED: Fibroblast growth factor 23(FGF23) is a bone-derived hormone which regulates mineral homeostasis but may also have a role in cardiovascular disease. Here, we found that higher plasma FGF23 was independently associated with decreased heart rate variability in stage 5 CKD patients and parathyroidectomy may reverse these abnormal indicators. INTRODUCTION: Lower heart rate variability (HRV) in patients with chronic kidney disease (CKD) compared with healthy controls is associated with increased risk of cardiovascular disease (CVD). Higher levels of plasma FGF23 also predict higher risk of CVD. Here, we aimed to evaluate the relationship between plasma FGF23 levels and HRV in patients with stage 5 CKD and to investigate longitudinal changes of them together with the correlation between their changes in two severe secondary hyperparathyroidism (SHPT) subgroups with successful parathyroidectomy (PTX) and persistent SHPT. METHODS: This cross-sectional study included 100 stage 5 CKD patients, 78 controls, and a prospective study in two PTX subgroups classified as successful PTX (n = 24) and persistent SHPT (n = 4) follow-up. Blood examination and 24-h Holter monitoring for HRV were measured. RESULTS: Most HRV indices were lower in stage 5 CKD patients than in healthy controls, and plasma FGF23 levels were higher. In multivariate stepwise regression models, levels of plasma FGF23 and serum parathyroid hormone (PTH) were correlated with HRV. The successful PTX subgroup had significant improvements over baseline in HRV indices. Persistent SHPT subgroup had numerically similar changes in HRV indices. However, plasma FGF23 levels decreased in both subgroups. CONCLUSIONS: Plasma FGF23 levels were higher in CKD patients than in controls, much higher in patients with severe SHPT. FGF23 was independently associated with decreased HRV in stage 5 CKD. Successful PTX may reverse these abnormal indicators and contribute to decreases in the risk of cardiovascular disease.

[Comparative analysis of the role of CD4(+) and CD8(+) T cells in severe asthma development].

The role of CD8^(+) T cells in asthma has not been fully discussed. The mechanisms of CD4^(+) and CD8^(+) cells in severe asthma (SA) development were compared. The microarray data (GSE31773) was downloaded from the Gene Expression Omnibus (GEO) database, including 20 samples of CD4^(+) and CD8^(+) T cells, which were collected from 8 health controls (HC), 4 non-severe asthma (NSA) and 8 SA patients. DEGs of CD4^(+) and CD8^(+) T cells in the HC vs. NSA and HC vs. SA groups were identified using the limma package in R. GO and pathway enrichment analysis of the common DEGs between the two groups were analyzed using DAVID. The interactive network of DEGs and significant modules were further explored. In CD4^(+) cells, there were 168 DEGs in HC vs. NSA group and 685 DEGs in HC vs. SA group, while for CD8^(+) T cells there were 719 DEGs in the HC vs. NSA groups and 1255 DEGs in the HC vs. SA groups. Besides, 80 common DEGs from CD4^(+) samples were enriched in the MAPKKK cascade and molecular metabolism, and 385 common DEGs of CD8^(+) T cells were significantly related with cell apoptosis and transformation. Moreover, two significant modules of DEGs in CD4^(+) were found to be involved with MPO and BPI. One module of CD8^(+) T cells containing PDHA1 and MRPL42 was identified to be related with glycolysis. In conclusion, MPO and BPI in CD4^(+), and PDHA1 and MRPL42 in CD8^(+) T cells might be used as specific biomarkers of SA progression. Therapy targeting the functions of CD4^(+) and CD8^(+) T cells may provide a novel perspective for SA treatment.

Association of IGF1R polymorphisms with the development of HBV-related hepatocellular carcinoma.

Although the involvement of insulin-like signaling in cancer has been well documented in various types of cancers, the association between the genetic variants in the insulin-like signaling and the development of hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC) remains unclear. In this study, a total of 498 individuals including 173 HBV related cirrhosis patients, 171 HBV-related HCC patients, and 154 healthy controls were enrolled. Sixteen single nucleotide polymorphisms (SNPs) in IGF1, IGF2, IGF1R and IGF2R have been genotyped by employing SNaPshot assays. We found A/A genotype at rs3743251 of IGF1R was negatively associated with HBV related HCC [odds ratio (OR) = 0.38, 95% confidence interval (CI) = 0.20-0.72, P = 0.037]; A/G genotype decreased the risk of portal vein thrombosis (OR = 0.38, 95%CI = 0.18-0.82, P = 0.01). These results indicate that rs3743251 polymorphism in IGF1R is associated with the susceptibility of HBV-related HCC.

Total parathyroidectomy with autotransplantation for a rare disease derived from uremic secondary hyperparathyroidism, the uremic leontiasis ossea.

SUMMARY: We described six uremic leontiasis ossea (ULO) patients who underwent total parathyroidectomy with autotransplantation. ULO demonstrated more a systemic disease than a simple craniofacial deformation. The surgery seemed an effective treatment to alleviate secondary hyperparathyroidism and to improve patients' quality of life. ULO may have a high postoperative recurrence tendency. INTRODUCTION: ULO is a rare disease derived from uremic secondary hyperparathyroidism (SHPT). Previous studies mostly focused on the craniofacial deformations. This study aims to investigate the systemic features of the disease and the surgical outcomes. METHODS: The present study retrospectively assessed six ULO patients who underwent total parathyroidectomy (TPTX) with autotransplantation (AT). Follow-up data were recorded. The follow-up status was considered as "effectiveness" if serum intact parathyroid hormone (iPTH) levels were <150 pg/mL in the first 3 days after surgery, or as "recurrence" if serum iPTH gradually increased >300 pg/mL during follow-up in patients whose status was initially considered as "effectiveness". RESULTS: Craniofacial deformations, short stature, thoracocyllosis, spine malformations, osteodynia, and muscle weakness were observed in all patients. Abnormal pulmonary functions were observed in five patients. After surgery, one patient died from respiratory failure. Surgery was effective in the remaining five patients with relieved osteodynia and stopped craniofacial deformation. A mean follow-up of 7.6 (4 to 12) months was available. Three patients suffered from recurrence of hyperparathyroidism originating from autografts. CONCLUSIONS: Our data suggests that ULO is not only a simple disease with craniofacial malformations but is a severe systemic disease leading to increased surgical risks. TPTX with AT seems an effective treatment to relieve SHPT and to improve quality of life. ULO may have a high postoperative recurrence tendency.

Value of diffusion weighted imaging in the differential diagnosis of benign and malignant breast lesions at 3.0T MRI.

The aim of this study was to investigate the correlation between apparent diffusion coefficients (ADCs), the relative apparent diffusion coefficient (rADC), and the pathological prognostic factor human epidermal growth factor receptor 2 (HER-2) in patients with breast cancer. A total of 64 women with breast cancer underwent breast diffusion-weighted imaging. HER-2 expression was detected in histological specimens. The ADC value, rADC value, and HER-2 level were determined. The ADC and rADC values of the breast cancer group were 1.1495 ± 0.1499 x 10(-3) and 0.6602 ± 0.0853, respectively. The differences in the ADC and rADC values between the two groups were statistically significant. There was no correlation between the ADC value and HER-2 expression in patients with breast cancer (r = -0.508, P = 0.043). However, the rADC value eliminated the individual differences to some extent. Therefore, compared to the ADC value, the rADC value had a better correlation with HER-2 expression.

Development of an improved rat model of dual graft liver transplantation with long-term survival.

Dual graft liver transplantation has been demonstrated to be feasible as well as effective in increasing the donor pool and in preventing the potential for small-for-size syndrome. However, little is known about the pathophysiological and immune processes following dual graft liver transplantation due to the lack of appropriate animal models. The aim of this study, therefore, was to establish an improved rat model of dual graft liver transplantation, with long-term survival. Male inbred rats were used as both donors and recipients. One middle lobe together with another right middle lobe from the livers of two different donors were used as the dual grafts. The "basin-shaped anastomosis" technique was used to connect the suprahepatic inferior vena cava; "Y-shaped bridge" and "three-cuff" techniques were adopted for the anastomosis of the portal veins; and the "two-stent" technique was used for the anastomosis of the bile ducts. Six of the ten recipients survived for more than 100 days after dual graft liver transplantation. There was no difference in graft survival between dual and whole liver transplantation. The long-term survivors with dual grafts from two different donors had unobstructed portal vein flow, unobstructed biliary tract dilatation, normal graft function, and well-preserved hepatic structure. Therefore, this improved model will be potentially useful for evaluating the pathophysiological processes, immune responses between dual grafts and recipient, and mechanisms underlying the liver regeneration in dual grafts after dual graft liver transplantation.

Effects of coupled plasma filtration adsorption on immune function of patients with multiple organ dysfunction syndrome.

OBJECTIVE: To investigate the effects of coupled plasma filtration adsorption (CPFA) on the immune function of patients with multiple organ dysfunction syndrome (MODS). METHODS: This study was a prospective, pilot, before-and-after self-crossover, clinical trial. Seven patients diagnosed with MODS and severe infection were randomly allocated to both 10 hours of CPFA and 10 hours of high-volume hemofiltration (HVHF) with a 12-hour interval and in random order. Serum concentrations of 7 cytokines including tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), interleukin-1beta (IL-1beta), interleukin-10 (IL-10), interleukin-1 receptor antagonist (IL-1Ra), and soluble tumor necrosis factor receptors 1 and 2 (sTNFR1 and sTNFR2) were measured during each treatment. The HLA-DR expression by the blood monocytes and the TNF-alpha production by the patients' blood (both spontaneous and lipopolysaccharide stimulated) were tested before and after the treatment. TNF-alpha production of normal human monocytes (THP-1 cells) incubated in vitro with the patient plasma was also measured. RESULTS: During CPFA, the fall in serum TNF-alpha and rise in serum IL-1Ra coincided with the rise in ratios of sTNFR2/TNF-alpha and IL-1Ra/IL-1beta (p<0.05), which were different from those seen within HVHF (p<0.05). HLA-DR expression increased after CPFA (84.32%-/+4.63% vs. 73.65%-/+11.52%, p=0.037), but there was no change after HVHF (p>0.05). Spontaneous and lipopolysaccharide-induced TNF-alpha production increased over time with CPFA (p=0.038, p=0.034, respectively), but did not change with HVHF (p>0.05). Patient plasma suppressed the production of TNF-alpha by cultured normal monocytes. This effect decreased over time with CPFA (p=0.041), but there was no effect with HVHF (p>0.05). CONCLUSIONS: CPFA was superior to HVHF in increasing the ratios of antiinflammatory to proinflammatory mediators, improving antigen presentation ability, and restoring leukocyte responsiveness. These findings suggest a potential role for CPFA in the treatment of MODS.

Therapeutic dosage of ozone inhibits autophagy and apoptosis of nerve roots in a chemically induced radiculoneuritis rat model.

OBJECTIVE: Radiculoneuritis characterizes by the neurogenic pain along the back of patients. This study aims to investigate the therapeutic effects of ozone on radiculoneuritis and the associated mechanisms in rat models. MATERIALS AND METHODS: A chemical radiculoneuritis rat model was successfully established. The rats were divided into 3 groups, including radiculoneuritis Model rats group (Model group, n=18), Ozone therapy group (n=18), and Normal control group (n=18). Ozone was administered at a dosage of 1 mg/kg/day. The electron microscope was used to observe autophagosomes in the cytoplasm. Immunohistochemistry assay was performed to examine cleaved caspase 3 and double-labeled immunofluorescence assay was used to detect light chain 3B (LC3B) and neuronal nuclear antigen (NeuN) expression. Quantitative Real-time PCR (RT-PCR) and Western blot were employed to evaluate the expression of LC3B, Beclin 1, phosphodiesterase 2A (PDE2A), and nuclear factor-kB p65 (NF-kBp65). RESULTS: Ozone significantly decreased autophagosomes formation and inhibited autophagy of nerve root cells in radiculoneuritis rat model. Ozone significantly decreased levels of autophagosomes initiator, LC3B, compared to Model group (p<0.05). Ozone significantly decreased cleaved caspase 3 expressions and alleviated apoptosis of nerve root cells compared to that of Model group (p<0.05). According to RT-PCR and Western blot assay, ozone significantly suppressed LC3B and Beclin 1 expression compared to that of Model group (p<0.05). Ozone significantly decreased PDE2A and NF-kB p65 expression compared to that of the Model group (p<0.05). CONCLUSIONS: Therapeutic dosage of ozone inhibits autophagy by suppressing LC3B and Beclin 1 expression and reduces apoptosis by blocking NF-kB signaling pathway.

Direct effects of dexamethasone on human podocytes.

Glucocorticoids are widely used in the treatment of human glomerular diseases, but their mode of action is poorly understood particularly in steroid-sensitive nephrotic syndrome, which is most common in childhood and is characterized by a lack of inflammation in the kidney. The podocyte is a key cell in the glomerulus in health and disease: until recently, human podocytes have been difficult to study in vitro. We have developed a conditionally immortalized human podocyte cell line transfected with a temperature-sensitive simian virus 40 transgene: when the transgene is inactivated in vitro, these cells adopt the phenotype of differentiated podocytes. We have used these cells to evaluate, using immunocytochemistry, reverse transcriptase-polymerase chain reaction, and Western blotting, direct effects of the glucocorticoid dexamethasone at concentrations designed to mimic in vivo therapeutic corticosteroid levels. Dexamethasone upregulated expression of nephrin and tubulin-alpha, and downregulated vascular endothelial growth factor. Effects on cell cycle were complex with downregulation of cyclin kinase inhibitor p21 and augmentation of podocyte survival, without any effect on apoptosis. We report cytokine production by human podocytes, especially interleukin (IL)-6 and -8; IL-6 expression was suppressed by dexamethasone. These potent direct effects on podocytes illustrate a novel mode of action of glucocorticoids and suggest potential new therapeutic strategies for glomerular disease.

VEGF-C promotes survival in podocytes.

Vascular endothelial growth factor (VEGF)-A is an autocrine survival factor for podocytes, which express two VEGF receptors, VEGF-R1 and VEGF-R3. As VEGF-A is not a known ligand for VEGF-R3, the aim of this investigation was to examine whether VEGF-C, a known ligand for VEGF-R3, served a function in podocyte biology and whether this was VEGF-R3 dependent. VEGF-C protein expression was localized to podocytes in contrast to VEGF-D, which was expressed in parietal epithelial cells. Intracellular calcium ([Ca2+]i) experiments demonstrated that VEGF-C induced a 0.74+/-0.09-fold reduction in [Ca2+]i compared with baseline in human conditionally immortalized podocytes (hCIPs; P<0.05, one sample t-test, n=8). Cytotoxicity experiments revealed that in hCIPs VEGF-C reduced cytotoxicity to 81.4+/-1.9% of serum-starved conditions (P<0.001, paired t-test, n=16), similar to VEGF-A (82.8+/-4.5% of serum-starved conditions, P<0.05, paired t-test). MAZ51 (a VEGF-R3 kinase inhibitor) inhibited the VEGF-C-induced reduction in cytotoxicity (106.2+/-2.1% of serum-starved conditions), whereas MAZ51 by itself had no cytotoxic effects on hCIPs. VEGF-C was also shown to induce a 0.5+/-0.13-fold reduction in levels of MAPK phosphorylation compared with VEGF-A and VEGF-A-Mab treatment (P<0.05, ANOVA, n=4), yet had no effect on Akt phosphorylation. Surprisingly, immunoprecipitation studies detected no VEGF-C-induced autophosphorylation of VEGF-R3 in hCIPs but did so in HMVECs. Moreover, SU-5416, a tyrosine kinase inhibitor, blocked the VEGF-C-induced reduction in cytotoxicity (106+/-2.8% of serum-starved conditions) at concentrations specific for VEGF-R1. Together, these results suggest for the first time that VEGF-C acts in an autocrine manner in cultured podocytes to promote survival, although the receptor or receptor complex activated has yet to be elucidated.

Expression of neuropilin-1 by human glomerular epithelial cells in vitro and in vivo.

Vascular endothelial growth factor (VEGF) is a potent promoter of endothelial mitogenesis and of endothelial permeability. Within the kidney it is synthesized primarily in the visceral glomerular epithelial cells (vGECs); however, the role of VEGF in the glomerulus remains unknown, as does the target cell upon which it acts. Although the target cells may be those of the glomerular endothelium, there are micro-anatomical reasons why this might not be the case. This, therefore, led us to consider the possibility that glomerular VEGF may bind to the vGECs themselves. Since it has been shown that vGECs do not express the main tyrosine kinase VEGF receptors, we chose to study vGEC expression of the more recently described VEGF isoform-specific receptors, the neuropilins. The expression of mRNAs for neuropilin-1, neuropilin-2 and soluble neuropilin was studied in whole kidney, sieved glomeruli and cultured podocytes by reverse transcription-PCR, and neuropilin-1 mRNA expression in isolated single glomeruli was analysed by nested reverse transcription-PCR. The expression of neuropilin-1 protein was investigated in cultured vGECs by Western blotting and immunocytochemistry, and in normal kidney sections by immunohistochemistry. Neuropilin-1 mRNA was detected in whole kidney, single and sieved glomeruli and cultured vGECs. Neuropilin-1 protein was detected in cultured vGECs and in vGECs in normal kidney sections by immunohistochemistry. Thus the present study suggests that vGECs may have the potential to bind the VEGF that they secrete. Functional studies will be required to address the potential significance of this finding in terms of an autocrine loop or VEGF sequestration.